The central unifying theme of this competing renewal is the sub- endothelial extracellular matrix (ECM) and its role in thrombus formation and dissolution. The collaborative efforts of four senior scientists will focus on the influence of cell adhesion, integrins, flow forces, proteases and protease inhibitors on the assembly and structure of ECM. Dr. Loskutoff will study interactions between plasminogen activator inhibitor 1 (PAI-1), vitronectin (VN), and the urokinase receptor (uPAR). Their influence on ECM structure and function, on the adhesion, migration, and growth of cells in vitro, and on tumor cell-mediated angiogenesis in vivo, will be defined. The origin of platelet VN will be examined, and its contribution to the composition of ECM and thrombi will be studied using VM- and PAI-1-deficient mice. Dr. Ginsberg will study intracellular mechanisms that control integrin functions essential for the assembly insoluble fibronectin fibrils in ECM. He will employ noel genetic strategies and structural mimics to define the mechanisms responsible for "activation" of the high affinity form of integrin alpha5beta1, and for its physical linkage to the cytoskeleton. Interactive sites in the integrin and the cytoskeletal proteins will be mapped, and the hypothesis that the differential binding of cytoskeletal proteins to integrins leads to integrin-specific functions will be tested. Dr. Ruggeri will study mechanisms responsible for platelet deposition at sites of vascular injury. He will define ECM components that influence platelet deposition at sites of vascular injury. He will define ECM components that influence platelet thrombus formation, characterize the thrombogenic properties of ECM proteins secreted by endothelial cells of different tissue origin and investigate the role of ECM in the bleeding defect in thrombospondin-2 deficient mice. The distinct roles of fibrinogen, fibronectin, vitronectin and integrin alphavbeta3 in platelet thrombus formation at normal and pathological shear rates will be assessed. Dr. Quigley will study mechanisms that initiate complex proteolytic enzyme cascades involved in ECM remodeling. The initiation and control of the uPA/plasmin cascade by a novel auto-activation step, by changes in intrinsic activity, and by PAI-1 and PAI-2 will be examined, and the link between this cascade and activation of select matrix metalloprotease cascades (MMP-3, MMP-9) will be investigated. Biochemical, cell biological and in vivo approaches using uPA-deficient and MMP-3- deficient mice will be employed. The work on the uPA/plasmin cascade is currently funded by NIH and will be folded into Project 4. The cores will be used by each of the projects to provide administrative activities and specialized monoclonal antibodies and to guarantee access to technically difficult in vitro and in vivo models of thrombus formation and dissolution.